CORE C. ADVANCED IMAGING CORE The overall goal of this program project grant is to study the mucosal immune barrier in infection and inflammation. This Core provides a facility for Advanced Imaging. Mucosal surfaces are largely lined by a monolayer of epithelial cells. Underlying the epithelial layer is extracellular matrix (ECM), connective tissue and vasculature. Inflammatory cells move bidirectionally from the blood across the endothelial cells, through the connective tissue and across the epithelial layer. We need microscopes that are capable of imaging all of these structures and processes. We must be able to image 3 dimensionally over time, i.e. 4 dimensional imaging. The microscopes and associated equipment must be capable of imaging both cultures (such as 3D cultures in ECM gels) and in vivo preparations. Cells and in vivo preparations must be kept alive and without photo-toxicity or other damage for extended periods. This Core contains 4 specialized microscopes at 2 locations. The Mission Bay facility has a Zeiss 510 confocal equipped with the META system and a Coherent Chameleon laser, as well as an environmental chamber. There is also a Zeiss Pascal confocal microscope equipped for photoactivation of GFP. The Parnassus facility contains a unique, custom built Robotic Spinning Disk Confocal Microscope for Intravital Timelapse imaging. It has already been used extensively for imaging cellular dynamics inside mammary glands in living, anesthetized mice. It has high performance temperature and environmental controls and a life support system for long term (150 hr) experiments. The microscope captures multi-color video rate images with negligible photobleaching. It is automated to repetitively image multiple sites and is equipped with state-of-the-art software. There is also an automated, time-lapse brightfield microscope for parallel monitoring of up to 96 wells (or any similar format) for as long as 200 hrs. This is particularly useful for screening of multiple conditions. Most infectious agents enter the body through exposed mucosal surfaces. These surfaces are mainly lined by a single layer of epithelial cells. We are studying how this epithelial barrier protects us against infection.